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. 2022 Jul 17;27(14):4560.
doi: 10.3390/molecules27144560.

Role of the Hydroxyl Radical-Generating System in the Estimation of the Antioxidant Activity of Plant Extracts by Electron Paramagnetic Resonance (EPR)

Daniele Sanna  1 , Angela Fadda  2
Affiliations

Role of the Hydroxyl Radical-Generating System in the Estimation of the Antioxidant Activity of Plant Extracts by Electron Paramagnetic Resonance (EPR)

Daniele Sanna et al. Molecules. .

Abstract

The scavenging activity of hydroxyl radicals, produced by the Fenton reaction, is commonly used to quantify the antioxidant capacity of plant extracts. In this study, three Fenton systems (Fe/phosphate buffer, Fe/quinolinic acid and Fe/phosphate buffer/quinolinic acid) and the thermal degradation of peroxydisulfate were used to produce hydroxyl radicals; the hydroxyl radical scavenging activity of plant extracts (ginger, blueberry juices and green tea infusion) and chemical compounds (EGCG and GA) was estimated by spin trapping with DMPO (5,5-dimethyl-1-pyrroline N-oxide) and EPR (Electron Paramagnetic Resonance) spectroscopy. Phosphate buffer was used to mimic the physiological pH of cellular systems, while quinolinic acid (pyridine-2,3-dicarboxylic acid) facilitates the experimental procedure by hindering the spontaneous oxidation of Fe(II). The EC50 (the concentration of chemical compounds or plant extracts which halves the intensity of the DMPO-OH adduct) values were determined in all the systems. The results show that, for both the chemical compounds and the plant extracts, there is not a well-defined order for the EC50 values determined in the four hydroxyl radical generating systems. The interactions of phosphate buffer and quinolinic acid with the antioxidants and with potential iron-coordinating ligands present in the plant extracts can justify the observed differences.

Keywords: DMPO; EPR; Fenton reaction; hydroxyl radical; plant extracts.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
A typical EPR spectrum of the DMPO–OH adduct.
Figure 2
Figure 2
Number of hydroxyl radicals, measured as DMPO–OH signal intensity (a.u. arbitrary units) and produced with the four radical-generating systems. Fe(II)/Bfr: Fe(II)-phosphate buffer; Fe(II)/Quin: Fe(II)-quinolinic acid; Fe(II)/Quin/Bfr: Fe(II)-quinolinic acid-phosphate buffer; Persulfate: thermal degradation of peroxydisulfate. Bars marked by asterisks differ significantly by Tukey’s test (p ≤ 0.05 *).
Figure 3
Figure 3
Percentage of inhibition as a function of EGCG concentration (A) and log EGCG concentration (B) in the Fe(II)/phosphate buffer system.
Figure 4
Figure 4
Graphical representation of EC50 and the corresponding CI 95% values for each plant juice, infusion and chemical compound. Each point represents a hydroxyl radical-generating system: Fe(II)/phosphate buffer (●), Fe(II)/Quin/phosphate buffer (●), Fe(II)/Quin (●), thermal degradation of peroxydisulfate (●).
Figure 5
Figure 5
Percentage of inhibition as a function of the EGCG concentration measured in the system Fe(II)/Quin.
See this image and copyright information in PMC

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